CAJ | 학술논문

为了异源合成抗疟疾药物青蒿素重要前体青蒿二烯，以酿酒酵母作为底盘细胞，利用基因工程手段构建功能人工酵母细胞。为提高基因拷贝数，并增加重组菌株基因型的稳定性，选择酵母基因组中多拷贝位点 Delta 为整合点，实现酿酒酵母内源基因tHMGR和ERG20的过表达以及外源基因ADS的整合。过表达tHMGR和ERG20基因增加了酵母体内半萜类物质共同前体法尼基焦磷酸FPP的积累量；而导入外源基因ADS，实现了酵母生产青蒿二烯。经过摇瓶发酵优化实验，人工酵母菌株青蒿二烯产量为225.3 mg·L?1；为了进一步提高青蒿二烯产量，经过发酵过程优化和补料策略，人工酵母菌株在5 L发酵罐中青蒿二烯产量达到1.05 g·L?1。
위료이원합성항학질약물청호소중요전체청호이희，이양주효모작위저반세포，이용기인공정수단구건공능인공효모세포。위제고기인고패수，병증가중조균주기인형적은정성，선택효모기인조중다고패위점 Delta 위정합점，실현양주효모내원기인tHMGR화ERG20적과표체이급외원기인ADS적정합。과표체tHMGR화ERG20기인증가료효모체내반첩류물질공동전체법니기초린산FPP적적루량；이도입외원기인ADS，실현료효모생산청호이희。경과요병발효우화실험，인공효모균주청호이희산량위225.3 mg·L?1；위료진일보제고청호이희산량，경과발효과정우화화보료책략，인공효모균주재5 L발효관중청호이희산량체도1.05 g·L?1。
Amorphadiene, the role precursor of anti-malaria drug artemisinin, was produced by artificial Saccharomyces cerevisiae cells that were constructed under guidance of gene engineering methods. Homologous genes tHMGR and ERG20 as well as heterogenous gene ADS were integrated into yeast genome at the multiple copies Delta sites. Two advantages of this strategy were to increase copy number of genes and to enhance the stability of recombinant strains genotype. Overexpression of tHMGR and ERG20 genes increased accumulation of FPP, the common precursor for sesquiterpenoids. The artificial yeast strains could produce amorphadiene by introducing heterogenous gene ADS. The production of amorphadiene was increased to 225.3 mg·L?1 in 50 ml flask fermentation through optimizing fermentation condition. In 5 L fermentor, production of amorphadiene was increased to 1.05 g·L?1 after optimization of fermentation process and the fed-batch feeding strategy.